The QIAexpress Type IV Kit is intended for molecular biology applications. This product is not intended for the diagnosis, prevention, or treatment of a disease.

pQE Vectors.

Elements listed in table

Performance

The QIAexpress Type IV Kit gives high expression, up to 50% of total cellular protein.

Principle

QIAexpresspQE vectors combine a powerful phage T5 promoter (recognized by E. coli RNA polymerase) with a double lac operator repression module to provide tightly regulated, high-level expression of recombinant proteins in E. coli. Protein synthesis is effectively blocked in the presence of high levels of lac repressor and the stability of cytotoxic constructs is enhanced. The pQE vectors (see figure pQE Vectors and table) enable placement of the His tag at either the N- or C-terminus of the recombinant protein.

Elements present in QIAexpress pQE Vectors

Element

Description

1. Optimized promoter/operator element

Consists of the phage T5 promoter and two lac operator sequences, which increase the probability of lac repressor binding and ensure efficient repression of the powerful T5 promoter

2. Synthetic ribosomal binding site RBSII

For efficient translation

3. His-tag coding sequence

Either 5' or 3' to the polylinker cloning region

4. Translational stop codons

In all reading frames for convenient preparation of expression constructs

5. Two strong transcriptional terminators

t0 from phage lambda, and T1 from the rrnB operon of E. coli, to prevent read-through transcription and ensure stability of the expression construct

6. ColE1 origin of replication

From pBR322

7. beta-lactamase gene (bla)

Confers ampicillin resistance

Procedure

Inserts encoding proteins of interest are cloned into appropriate constructs and transformed into a suitable E. colistrain for expression. Expression is induced by addition of IPTG. QIAexpress Type IV Kit constructs can be transformed into E. coli, used as a shuttle vector for recombinant protein expression in insect cells, or transfected into mammalian cells.

Applications

The QIAexpress Expression System provides high-level expression of proteins suitable for many applications, including: